USPTO Patents Application 10687850

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EuropSlsches Patentamt
European Patent Office
Office europeen des brevets

111

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(ID EP1 129 211 B1

EUROPEAN PATENT SPECIFICATION

(45) Date o{ publication and mention
of the grant of the patent:
09.07.2003 Bulletin 2003/28

(21) Application number. 99954225.1

(22) Date of filing: 11.11.1999

(51) Intci7: C12Q 1/00, G01N 35/00,
G01 N 27/30, G01N 27/327

(86) International application number:
PCT/GB99/03764

(87) International publication number:

WO 007028068 (18.05.2000 Gazette 2000/20)

(54) ELECTRODE STRIPS FOR TESTING SMALL VOLUMES

ELEKTRODENSTREI FEN ZUM TESTEN VON KLEINEN VOLUMEN
BANDES ELECTRODES PERMETTANT DE TESTER DES PETITS VOLUMES

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(84) Designated Contracting States:

AT BE CH CY DE DK ES Fl FR GB GR IE IT LI LU
MCNLPTSE

(30) Priority: 11.11.1998 GB 9824627

(43) Date of publication of application:
05.09.2001 Bulletin 2001/36

(73) Proprietor: Cambridge Sensors Ltd.

Godmanchester, Cambs PE29 2XG (GB)

(72) Inventors:

• YON HIN, Bernadette
Cambridge Sensors Limited
Cambridge CB4 1XT (GB)

• MCCANN, James Cambridge Sensors Limited
Cambridge CB4 1XT (GB)

• BLAIR, Nell

Hardwick Cambridge CB3 7XU (GB)

• COX, Lorna, Jean Cambridge Sensors Limited
Cambridge CB41XT(GB)

(74) Representative: Perry, Robert Edward
GILL JENNINGS & EVERY
Broadgate House
7 Eldon Street
London EC2M 7LH (GB)

(56)

References cited:
EP-A- 0 230 472
WO-A-97/02487
US-A-4 218 421
US-A- 5 628 890
US-A- 5 779 867

EP-A- 0 593 096
WO-A-99/13100
US-A- 5 169 600
US-A- 5 679 311
US-A- 5 798 031

Note: Within nine months from the publication of the mention of the grant of the European patent, any person may give
notice to the European Patent Office of opposition to the European patent granted. Notice of opposition shall be filed in
a written reasoned statement. It shall not be deemed to have been filed until the opposition fee has been paid. (Art.
99(1) European Patent Convention).

Printed by Jouve, 75001 PARIS (FR)

Description
I0001J This invention relate tn

^•^^-.CiltS* s,rips for

EP 1 129 211 B1

cation of the reagents ^S^* B ;««*

C eXPerie " Ced * users o,

trodes which LState fhi * Pr ° Ximity ,0 0,6 etec "
5141 868 EP-A-01 70375 and US-A-

Iv small to. . ^ Wh0se tensions are sufficient-
V small lo allow introduction of a sample e a pV? ,
in volume, by capillarv action Th« T 9 " 5-3 »* L

ed on the octto criecfrode mediator that are coat-

mined. The active 2 he blood can be ^ter-
samplewithoutanmte^n 6 ' S exposedt °a "hole blood
bloodfilter " en,n9membraneorotn ^whole

rnonofHame^tlSove^lT CtiVe,ay ^ anda
ence electmHo » BW i 0Wer »'e reagent and the refer-

S^scnptipno^

§iB ngryofthe Invent?™

SSLSSSK** r sem invention - a «•* ^

brane wimlnwhich a sma r? ament mesh or me ">
can be 522 21 lvol " meofHt >"W to be tested

electrodes an J wherl Pr ° V,de ^ betwee " the

suchasone:„:n:r" a " ana, ^- s p^ reagent

*vme. co-fac,orTS a , 0 r IT Tf 0 ^ e * M en "
membrane The teswr , ' C ° a,ed on ,he mes » or
he test strip also includes a sample appli-

ticulady indioa e"es wCr nfi9Ura,i ° n ' S par "
tow. the sample is vZZ , V * SBmp ' e vo,um ° *
u •* a rapid -^ESS? 98 *'< b ^ «*o->

-^SE£5£ ™ e r °' the workin 9

2

3

EP1 129 211 B1

4

aqueous solution by pipetting. Afurther layer containing
NAD is then deposited onto the working electrode.
[001 2J A monofilament mesh material is coated with
a surfactant and then with a solution containing glucose
dehydrogenase via pipetting, ink jet-coating or dip-coat- 5
ing, and is placed over the two electrodes to form a re-
action chamber. This reaction chamber may be defined
further by additional printing, or by the use of a top layer
to form an edge fill cavity. For example, a second non-
conducting ink printed on top of the mesh material, and 10
then a cover tape is applied on top of the mesh in such
a way as to leave an extended area of the mesh exposed
for sample application.

[0013] The device allows the application of a small
volume of sample (typically 1 uX or less) to the mesh '5
extension. This is followed by flooding of the device
sensing area with sample, bringing it into intimate con-
tact with the measuring electrodes.
[0014] Devices having an edge fill are described In
WO-A-98/55856. They can be simply adapted, in ac- 20
cordance with the present invention. In particular, refer-
ence may be made to Fig. 1 in WO-A-98/55856; com-
ponents of this invention are the support (1), electrodes
(2/3), mesh material (6) and tape (7); in addition, reagent
is provided on the mesh material. Such a device can 25
work by application at its edge, to a sample. This is par-
ticularly valuable in cases where it is difficult to extract
the sample. Other configurations will be evident to one
skilled in the art, including combinations of one or more
of the cof actor, mediator or the enzyme coated onto the' 30
overlying mesh or membrane sheets. The choice of
combination may on the reaction kinetics of the various
compounds.

[0015] In another embodiment of the device, the en-
zyme or the mediator is coated on the sheet, the co- 35
factor and the other of the mediator or the enzyme are
coated onto the working electrode directly, and the sheet
is capable of filtering the whole blood such that the ac-
tive electrode sees a sample which is effectively free of
whole blood cells. In this case, the haematocrit depend- *o
ency of the result is substantially reduced. In this man-
ner, the cell-filtering function of a selected membrane
may be combined with the rapid kinetics of having the
some or ail of the active elements of the reaction (the
enzyme, mediator and the co-factor) in the membrane, *s
to produce a highly effective device.
[0016] In summary, according to the present inven-
tion, a device is constructed by depositing one or more
of the reagents required for the quantitation of an ana-
lyte as a single or multiple layers on a fine mesh material 50
or membrane; the deposited areas are of dimensions
small enough to wet with a very small sample volume.
The mesh ormembrane can be used in both colorimetric
and electrochemical devices.

[0017] A characteristic of this invention is that a rea- 55
gent is applied precisely onto a target area on a woven
material such as polyester or nylon or other porous
membrane. In use, this provides rapid solubilisation of

the reagents in the presence of the sample. The reagent
or reagents can be applied in a number of different meth-
ods that result in the deposition of a known volume at a
precise location and in a well-defined foot-print. These
include the use of dispensing equipment such as a pis-
ton pump, syringe pump or on-demand ink-jet printer.
[001 8] Also described is a flexible tape contain ing one
or more reagents may be laminated to another flexible
tape on which is printed a series of electrodes. Instead
of cutting out individual sensors, the laminate (compris-
ing a row or series of sensors) may be used sequentially,
e.g. on being dispensed from a suitable dispenser. For
this purpose, whether or not as a laminate, a tape of the
invention may be provided as a roll, and stored in sealed
cassettes which may also contain desiccant. In use, the
cassette may be inserted into a automatic dispenser
from which the tape, is wound out automatically by an
indexing mechanism to reveal sequentially the discrete
sensors. The action of this instrument is therefore anal-
ogous to the action of a film in a camera. In this embod-
iment, the tape may also contain a red blood cell-lysing
reagent such as saponin, in order to reduce the effect
of haematocrit and haemoglobin in a whole blood sam-
ple. The tape may be further protected from moisture by
being covered with a peelable film (e.g. of aluminium)
that is automatically peeled off when the tape is dis-
pensed from the cassette. When the sample is applied
to the sensor, the amount of analyte of interest in the
sample may be determined electrochemically. Such de-
termination can be conducted by known methods.
[0019] The following Example illustrates the inven-
tion.

Example

[0020] A conductive ink material is printed onto a non-
conducting polyester sheet material by a screen-printing
process. The conductive ink material consists of a mix-
ture of graphite and carbon particles and a polymer bind-
er in an organic solvent. After deposition of the conduc-
tive ink, solvents are removed in a forced air oven. A
silver/silver chloride reference/counter electrode is
printed onto one of each pair of printed carbon elec-
trodes followed by a non-conducting ink layer to define
the contact pads and the sensor area.
[0021 ] A mediator such as Meldola Blue, Nile Blue or
other suitable dye and the enzyme co-factor nicotina-
mide adenine dinucleotide (NAD) are deposited onto the
carbon electrode. Alternatively, the NAD is applied sep-
arately over the mediator from an aqueous ink.
[0022] The enzyme glucose dehydrogenase is de-
posited as uniform spots on a monofilament polyester
mesh tape. This is achieved as follows:

(a) in a contact mode, where a drop formed at a dis-
penser tip in close proximity to the mesh is allowed
to be transferred to the mesh by touching off the
drop onto the mesh surface; or

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EP 1 129 211 B1

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(b) in a non-contact mode, where a drop formed by
an ink-jet printhead or other orifice above the mesh
is dropped onto the mesh from a distance under
conditions which do not cause it to penetrate the
mesh.

[0023] Upon drying, the spots spread to cover an area
defined partly by the characteristics of the mesh weave
and partly by the application conditions. Typically the ar-
eas covered by a 500 nL drop is 1 .3 x 1 .2 mm. The mesh
tape is allowed to dry at room temperature.
[0024] The enzyme-modified mesh tape is then lami-
nated onto the modified sheet of devices and secured
further by a non-conducting print. Finally, a cover tape
is laminated on tope of the mesh. The sheets of devices
are disc cut into individual devices. In an alternative de-
vice format, the laminated sheets are wound and includ-
ed in a cassette type unit, allowing a single device to be
used by a wind-on mechanism similar to a camera film-
winding system.

Claims

1 . A test strip comprising a support carrying an active
electrode and a counterelectrode, and a monofila-
ment mesh or membrane within which a small vol-
ume of liquid to be tested can be distributed and
provide contact between the electrodes, wherein an
anaiyte-specific reagent is coated on the mesh or
membrane, and wherein the test strip includes a
sample application area at one edge of the mesh or
membrane.

2. A test strip according to claim 1 , wherein the rea-
gent is at least one component of a redox reaction.

3. A test strip according to claim 2, wherein the at least
one compound is one or more of an enzyme, a me-
diator and/or co-factor for the enzyme.

4. A test strip according to claim 2, wherein the at least
one component comprises an enzyme.

5. A test strip according to claim 3 or claim 4, wherein
the enzyme is glucose oxidase or glucose dehydro-
genase.

6. A method for testing a liquid for the presence of an
analyte, which comprises contacting the liquid with
a test strip according to any of claims 1 to 5, and
detecting the current.

7. A method according to claim 6, wherein the liquid
is blood and the analyte is glucose.

Patentansprtiche

1. Teststreifen, umfassend einen eine aktive Etektro-
de und eine Gegenelektrode tragenden Trager so-

5 wie ein Monofilament-Gewebe bzw. eine Membran,
worin ein kleines Volumen einer zu testenden Flus-
sigkeit verteilt werden und als Kontakt zwischen
den Elektroden dienen kann, wobei das Gewebe
bzw. die Membran mit einem analytspezifischen

10 Reagens beschichtet wird und wobei der Teststrei-
fen einen Probenauftragsbereich an einem Ende
des Gewebes bzw. der Membran enthalt.

2. Teststreifen nach Anspruch 1, wobei es sich bei
15 dem Reagens urn wenigstens eine Komponente ei-
ner Redoxreaktion handelt.

3. Teststreifen nach Anspruch 2, wobei es sich bei der
wenigstens einen Verbindung urn ein oder mehrere

20 Enzyme, einen Mediator und/oder einen Cofaktor
fur das Enzym handelt.

4. Teststreifen nach Anspruch 2, wobei die wenig-
stens eine Komponente ein Enzym umfaBt.

25

5. Teststreifen nach Anspruch 3 oder Anspruch 4, wo-
bei es sich bei dem Enzym urn Glucose-Oxidase
oder Glucose-Dehydrogenase handelt.

30 6. Verfahren zum Testen einer Flussigkeit auf die An-
wesenheit eines Analyten, wobei die Flussigkeit mit
einem Teststreifen nach einem der Anspruche 1 bis
5 in Kontakt gebracht und der Stromfluss nachge-
wiesen wird.

35

7. Verfahren nach Anspruch 6, wobei es sich bei der
Flussigkeit urn Blut und bei dem Analyten um Glu-
cose handelt.

40

Revendications

1. Bande d'essai comprenant un support portant une
electrode active et une contre-electrode, et un filet

45 monofilamentaireou membrane a I'interieurduquel
un faible volume de liquide a tester peut etre reparti
et assurer un contact entre les electrodes, dans la-
quelle un reactif particulier a un analyte est applique
sur le filet ou la membrane et dans laqueile la bande

so de test comprend une zone d'application d'echan-
tillon en un bord du filet ou de la membrane.

2. Bande d'essai selon la revendication 1 , dans laquei-
le le reactif est au moins un composant o"une reac-

55 tion d'oxydoreduction.

3. Bande d'essai selon la revendication 2, dans laquei-
le le au moins un composant est I'un ou plusieurs

EP1 129 211 B1

d'une enzyme, d'un mediateur et/ou un cofacteur
pour Tenzyme.

4. Bande d'essai selon la revendication 2, dans laquel-

le le au moins un composant comprend une enzy- s
me.

5. Bande d'essai selon la revendication 3 ou ia reven-
dication 4, dans laquelle I'enzyme est une oxydase

de glucose ou une deshydrogenase de glucose. io

6. Precede de test d'un liquide en presence d'un ana-
lyte, qui comprend la mise en contact du liquide
avec une bande d'essai selon Tune quelconque des
revendications 1 a 5, et a detecter le courant. is

7. Procede selon la revendication 6, dans lequel le li-
quide est le sang et Panalyte est le glucose.

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